Vishal Gaurav - Pharmaceutically-relevant diterpenoid biosynthesis: Assay development, optimization and quantification

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  Publication Details (including relevant citation   information):

  Authors: Vishal Gaurav, Susan C. Roberts

  Publication date: 2009

  Conference name: The American Chemical Society (ACS) Proceedings

  Publisher: The ACS National Meeting, Salt Lake City, UT


  Paclitaxel, the best-selling anti-cancer drug is a complex   diterpenoid synthesized in Taxus plant cell cultures through a   multi-step biosynthetic pathway. To unravel the inherent culture   heterogeneity and production variability, we are investigating   regulation of cellular metabolism through flow cytometric cell   sorting technologies. A novel antibody-based indirect   immunofluorescent assay was developed to detect paclitaxel   accumulation in live plant cells. Concomitantly, fluorescein   diacetate assay was performed to ensure cell viability.   Experiments were performed with variations in antibody   concentrations, wash steps and incubation times to establish an   optimal procedure. Subsequently, a novel flow cytometry-based   method to quantify the alkaloid content, by investigating the   antigenantibody interactions, was developed.

  Address (URL): iosynthesis_in_plant_cells.pdf